Construction method of plasma cell-free dna library

Abstract

The invention discloses a 'construction method for a small amount of DNA library'. The method comprises the following steps: carrying out end repairing on plasma free DNA or fragmented DNA and simultaneously carrying out phosphorylation modifying on 5' terminal to obtain modified DNA; later, implementing blunt end linking directly to an artificial linker and purifying to obtain relatively pure linker-containing DNA having a gap on joint; then repairing the gap by virtue of polymerase chain reaction (PCR) polymerase before PCR amplification to obtain repaired complete linker-containing DNA; and carrying out PCR amplification reaction and purifying a finished product to obtain a final high-throughput sequencing library. The method is not only simple, convenient and efficient, and is capable of avoiding a purification step and reducing DNA loss in a library construction process, but also can effectively solve a problem of linker self-linking which is easy to occur in a micro-DNA library construction process by virtue of a special linker and blunt end linking way; and meanwhile, by repairing the gap before amplification, sufficient complete templates are provided before amplification for the amplification reaction.

Description

technical field [0001] The invention relates to a method for constructing a plasma DNA library for gap repair before amplification, in particular to a method for constructing a free DNA library in plasma of a pregnant woman that can be applied to non-invasive prenatal diagnosis. Background technique [0002] According to medical statistics, about 2 to 3% of newborns born each year suffer from congenital diseases or birth defects, resulting in long-term sickness, disability, and even premature death of infants and young children, bringing huge burdens to society and families. The main causes of congenital diseases or birth defects are chromosomal abnormalities, single-gene genetic diseases, polygenic genetic diseases, teratogenic factors, etc., among which chromosomal abnormalities account for a large proportion. In the modern society that emphasizes eugenics and health care, how to prevent congenital diseases or birth defects has become a very important issue. Prenatal diag...

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a method for constructing a high-throughput sequencing library to solve the problems of DNA loss and self-connection of adapters in the process of building a small amount of DNA samples, especially in the process of building a plasma cell-free DNA library.

This method is not only simple and efficient, but also reduces purification steps and DNA loss during library construction, and uses specific adapters and blunt-end connection methods to effectively solve the problem of self-ligation of adapters during the construction of trace DNA libraries.

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